Excitatory synapses in the brain are characterized by a dense network of proteins called the postsynaptic density (PSD) which contains receptors, scaffold proteins, and signaling molecules. Four members of a family of scaffold proteins called membrane-associated guanylate kinases (MAGUKs) are abundantly expressed in the PSD: PSD-95, PSD-93, and SAP102. Importantly, MAGUKs associate with a host of signaling proteins which coalesce into large complexes associated with NMDA type glutamate receptors (NMDARs) through MAGUK interactions with the cyptoplasmic C-terminal tails of NMDAR NR2 subunits. In addition, NMDAR GluN2 (as well as GluN1) subunits also contain binding sites that directly recruit additional downstream signaling pathways into these NMDA receptor signaling complexes or NRSCs. Although the formation of NRSCs is thought to provide a mechanism for organizing and coordinating activation of downstream signaling pathways following NMDAR activation, the role of NRSCs in NMDAR signaling and synaptic plasticity is poorly understood. For example, the role of protein interactions mediated by NMDAR GluN2 subunits is highly controversial and little is known about how protein interactions dependent on NMDAR GluN1 subunits are involved in NRSC signaling and synaptic plasticity. In this project we propose to address both of these issues using a combination of electrophysiological, biochemical, and molecular genetic approaches to delineate the roles of protein interactions dependent on the C-termini of GluN1 and GluN2 subunits in NMDAR signaling and synaptic plasticity. In Specific Aims 1 and 2 we will examine hippocampal synaptic plasticity and NMDAR signaling in genetically engineered mice where the C-terminus of GluN2A subunits has been deleted by replacing it with the C-terminus of GluN2B subunits. Mutants with the opposite mutation - deletion of the GluN2B C-terminus by replacing it with the C-terminus of GluN2A subunits will also be examined. In Specific Aim 3 we will use a similar overall approach to examine the role of protein interactions mediated by NMDAR GluN1 subunits in NRSC signaling and synaptic plasticity. Here, we will perform electrophysiological studies of synaptic plasticity and biochemical studies of NMDAR signaling in mice with null mutations in the GluN1 subunit adaptor protein AKAP9 as well as the AKAP9-associated protein kinase TNiK. Importantly, several of the proteins to be examined in this proposal have been implicated in diseases such as mental retardation and schizophrenia. Thus, our studies will not only provide fundamental insights into the mechanisms underlying activity-dependent forms of synaptic plasticity involved in learning and memory, but may also provide important information about how alterations in NRSC function may contribute to cognitive disorders and psychiatric disease. PUBLIC HEALTH RELEVANCE: Understanding the molecular mechanisms underlying the storage of new information in the brain during memory formation is a crucial first step toward the development of novel treatments for memory disorders. In this project we will investigate how the interaction of multi-protein complexes with neurotransmitter receptors regulates changes in neuronal function thought to underlie memory formation. Importantly, several of the proteins within these complexes have recently been implicated in diseases such as mental retardation and schizophrenia and thus our findings will not only shed new light on the molecular mechanisms of memory formation but also provide new insights into how disruption of these complexes contributes to cognitive disorders and psychiatric disease.